Effects Of A Novel Camptothecin Analogue, ISO-camptothecin On Proliferation And Cell Cycle Of Tumor Cells, Venular Endothelial Cells And Keratinocytes

Objective: Camptothecins (CPTs) are DNA topoisomeraseⅠinhibitors which have been extensively used clinically. In the recent years, attentions have been focused on developing effective CPT analogues with lower toxicity. Studies have showed that CPTs could inhibit proliferation, induce apoptosis and promote differentiation in tumor cells, venular endothelial cells and keratinocytes. Iso-camptothecin (Iso-CPT) is a novel CPT analogue. In this study, human squamous carcinoma cell Tca8113, human umbilical venular endothelial cell ECV304 and human keratinocyte colo-16 are treated with Iso-CPT to observe its effects on proliferation and cell apoptosis, in relation to potential use in treating tumors and psoriasis .Methods: The cultured Tca8113, Hela, ECV304 and colo-16 cells were exposed to iso-CPT at different concentration for 24,48,72 hours, respectively. Inhibition on cells proliferation and valid concentration of iso-CPT were measured by MTT assay. Morphologic changes were observed by inverted microscope. The influence on cell cycle and induction of cell apoptosis of iso-CPT were evaluated by Flow cytometry. Early apoptotic cells were quantitively detected by Annexin-PI assay.Results: (1)The results showed that iso-CPT exerted a notable inhibitory effect on proliferation of Tca8113, Hela, ECV304 and colo-16 cells. After treated by iso-CPT for 24,48,72 hours, the IC50 of Tca8113 cell was 78.19,73.12,61.36μg/ml, respectively; the IC50 of Hela cell was 80.23,76.02,64.29μg/ml; the IC50 of ECV304 cell was 76.38,71.65,59.57μg/ml; the IC50 of colo-16 cell was 76.12,68.42,57.84μg/ml. The inhibition revealed time and dose dependence in effective concentration range. (2) After exposure to iso-CPT, cells presented morphologic changes, including cell shrinkage, cell rounding and cytoplasmic blebbing; and the cell numbers in unit area reduced. (3) Iso-CPT could influence cell cycle of Tca8113 and ECV304, arrested cells in S phase. This effect showed time and dose dependence. (4) Iso-CPT interfered with colo-16 cell cycle and arrested cells in S and G2 phase ,the rate of S and G2 phase raised parallel to the increase in concentration and treated time. Flow cytometry results demonstrated that iso-CPT induced colo-16 cell apoptosis at the concentration of 31.25 and 50μg/ml. The apoptotic rate was 1.3% and 4.9%, respectively. When colo-16 was treated with 50μg/ml iso-CPT for 24, 48hours, the apoptotic rate was 1.5% and 5.6%, respectively. (5) When colo-16 cells were incubated with 50μg/ml iso-CPT for 48 hours ,the early apoptotic rate was 4.45%. (6)The inhibitory concentrations of iso-CPT to ECV304 and colo-16 cell were lower than those to tumor cells Tca8113 and Hela.Conclusion: (1) Iso-CPT can inhibit the proliferation of Tca8113 and Hela cells and this effect has time and dose dependence. Iso-CPT can influence the cell cycle of Tca8113 and arrest the cells in S phase, suggesting that its antitumor mechanism is relevant to this effect. (2)Iso-CPT can inhibit ECV304 cells proliferation, influence the cell cycle of ECV304 and arrest the cells in S phase, indicating that Iso-CPT is worth further research as a potential antiangiogenic agent. (3) Iso-CPT can inhibit colo-16 cell proliferation, influence its cell cycle, arrest the cells in S and G2 phase and induce cell apoptosis. Iso-CPT may have therapeutic effect on psoriasis. (4)The inhibitory concentration of Iso-CPT to ECV304 and colo-16 cell are lower than that to tumor cell Tca8113, suggesting that relative lower dose Iso-CPT may have effect on the treatment of psoriasis .